Mutant RAS-driven tumorigenesis was thought for decades to arise independently of wild-type RAS isoforms, but recent evidence points to their involvement. In this issue of Cancer Cell, Grabocka et al. report how loss of wild-type RAS alters oncogenic signaling and dampens the DNA-damage response, thereby promoting tumor progression and chemosensitivity.
Recently, tomatoes have been implicated as a primary vehicle in food-borne outbreaks of Salmonella enterica serovar Newport and other Salmonella serovars. Long-term intervention measures to reduce Salmonella prevalence on tomatoes remain elusive for growing and postharvest environments. A naturally occurring bacterium identified by 16S rRNA gene sequencing as Paenibacillus alvei was isolated epiphytically from plants native to the Virginia Eastern Shore tomato-growing region. After initial antimicrobial activity screening against Salmonella and 10 other bacterial pathogens associated with the human food supply, strain TS-15 was further used to challenge an attenuated strain of S. Newport on inoculated fruits, leaves, and blossoms of tomato plants in an insect-screened high tunnel with a split-plot design. Survival of Salmonella after inoculation was measured for groups with and those without the antagonist at days 0, 1, 2, and 3 and either day 5 for blossoms or day 6 for fruits and leaves. Strain TS-15 exhibited broad-range antimicrobial activity against both major food-borne pathogens and major bacterial phytopathogens of tomato. After P. alvei strain TS-15 was applied onto the fruits, leaves, and blossoms of tomato plants, the concentration of S. Newport declined significantly (P ≤ 0.05) compared with controls. Astonishingly, >90% of the plants had no detectable levels of Salmonella by day 5 for blossoms. The naturally occurring antagonist strain TS-15 is highly effective in reducing the carriage of Salmonella Newport on whole tomato plants. The application of P. alvei strain TS-15 is a promising approach for reducing the risk of Salmonella contamination during tomato production.
Here we report the genome sequence of a Clostridium botulinum strain IBCA10-7060 producing botulinum neurotoxin serotype B and a new toxin serotype. Multilocus sequence typing analysis revealed that this strain belongs to a new sequence type, and whole-genome single nucleotide polymorphism analysis showed that this strain clustered with strains in lineage 2 from group I.
Genome stability is essential for neural development and the prevention of neurological disease. Here we determined how DNA damage signaling from dysfunctional telomeres affects neurogenesis. We found that telomere uncapping by Pot1a inactivation resulted in an Atm-dependent loss of cerebellar interneurons and granule neuron precursors in the mouse nervous system. The activation of Atm by Pot1a loss occurred in an Atr-dependent manner, revealing an Atr to Atm signaling axis in the nervous system after telomere dysfunction. In contrast to telomere lesions, Brca2 inactivation in neural progenitors also led to ablation of cerebellar interneurons, but this did not require Atm. These data reveal that neural cell loss after DNA damage selectively engages Atm signaling, highlighting how specific DNA lesions can dictate neuropathology arising in human neurodegenerative syndromes.
ATM; cerebellum; DNA damage; neural development; telomeres
Phage typing has been used for the epidemiological surveillance of Salmonella enterica serovar Enteritidis for over 2 decades. However, knowledge of the genetic and evolutionary relationships between phage types is very limited, making differences difficult to interpret. Here, single nucleotide polymorphisms (SNPs) identified from whole-genome comparisons were used to determine the relationships between some S. Enteritidis phage types (PTs) commonly associated with food-borne outbreaks in the United States. Emphasis was placed on the predominant phage types PT8, PT13a, and PT13 in North America. With >89,400 bp surveyed across 98 S. Enteritidis isolates representing 14 distinct phage types, 55 informative SNPs were discovered within 23 chromosomally anchored loci. To maximize the discriminatory and evolutionary partitioning of these highly homogeneous strains, sequences comprising informative SNPs were concatenated into a single combined data matrix and subjected to phylogenetic analysis. The resultant phylogeny allocated most S. Enteritidis isolates into two distinct clades (clades I and II) and four subclades. Synapomorphic (shared and derived) sets of SNPs capable of distinguishing individual clades/subclades were identified. However, individual phage types appeared to be evolutionarily disjunct when mapped to this phylogeny, suggesting that phage typing may not be valid for making phylogenetic inferences. Furthermore, the set of SNPs identified here represents useful genetic markers for strain differentiation of more clonal S. Enteritidis strains and provides core genotypic markers for future development of a SNP typing scheme with S. Enteritidis.
Salmonella enterica subsp. enterica serovar Cubana (Salmonella serovar Cubana) is associated with human and animal disease. Here, we used third-generation, single-molecule, real-time DNA sequencing to determine the first complete genome sequence of Salmonella serovar Cubana CFSAN002050, which was isolated from fresh alfalfa sprouts during a multistate outbreak in 2012.
Analyses used data from an extended longitudinal study to examine the relationship between childhood physical and sexual abuse (CPA and CSA, respectively) and adolescent and adult smoking behavior. Two questions guided the study: (1) Is there an association between childhood abuse and adolescent and adult smoking behavior? (2) Does the relationship between childhood abuse and later cigarette smoking differ for males and females?
A censored-inflated path model was used to assess the impact of child abuse on adolescent and adult lifetime smoking prevalence and smoking frequency. Gender differences in significant model paths were assessed using a multiple-group approach.
Results show no significant relation between CPA or CSA and risk of having ever smoked cigarettes in adolescence or adulthood. However, for males, both CPA and CSA had direct effects on adolescent smoking frequency. For females, only CSA predicted increased smoking frequency in adolescence. Adolescent smoking frequency predicted adult smoking frequency more strongly for females compared to males.
CPA and CSA are risk factors for higher frequency of smoking in adolescence. Higher frequency of cigarette smoking in adolescence increases the risk of higher smoking frequency in adulthood. Results underscore the need for both primary and secondary prevention and intervention efforts to reduce the likelihood of childhood abuse and to lessen risk for cigarette smoking among those who have been abused.
substance use; smoking; cigarettes; trauma; child abuse; sexual abuse; physical abuse
To examine evidence of the continuity in abusive discipline across two generations (G1 and G2) and the role of Safe, Stable, and Nurturing Relationships (SSNRs) as protective factors.
Data are from the Lehigh Longitudinal Study, a prospective investigation of the causes and consequences child maltreatment that began in the 1970s with a sample of 457 children and their parents. Data were most recently collected in 2008 - 2010 from 80% of the original child sample (N = 357) when they were adults age 36 years on average. Of those assessed as adults, 268 participants (G2s) were parenting children and thus comprise the analysis sample. Analyses examined the association between physical discipline practices by G1 parents and G2’s reports of similarly severe discipline practices used in parenting their own children. Analyses also investigated the direct and interactive (protective) effects of SSNR variables that pertain to the care, warmth, and support children received from their mothers, fathers, and siblings over their lifetimes. A measure of an adult partner’s warmth and support was also included. A case-level examination of G2 harsh discipliners was included to examine other forms of past and more recent forms of abuse exposure.
Results show a significant predictive association between physical discipline by G1 and G2 parents (Beta = .30, p < .05, OR: 1.14 [1.04 - 1.26]), after accounting for childhood socioeconomic status and gender. While being physically disciplined as a child was inversely related to reports of having had a caring relationship with one’s mother (r = -.25, p < .01), only care and support from one’s father predicted a lower risk of harsh physical discipline by G2s (Beta = -.24, p < .05, OR: .74 [.59 -.92]). None of the SSNR variables moderated the effect of G1 discipline on G2 discipline. A case-level examination of the abusive histories of G2 harsh discipliners found they had in some cases been exposed to physical and emotional abuse by multiple caregivers and by adult partners.
There is continuity in physical disciplining across two generations. SSNRs, while requiring further examination, did not mediate or moderate the effect of G1 on G2 harsh physical discipline. Care and support from fathers was inversely related to the likelihood of G2 harsh physical discipline, although this relationship is independent of abuse in childhood. Research is needed to identify factors that interrupt the continuity of abuse in two or more generations so that promising interventions can be implemented.
intergenerational abuse; protective factor; mitigating factor; safe; stable; nurturing; relationship (SSNR)
Mucus production by goblet cells of the large intestine serves as a crucial anti microbial protective mechanism at the interface between the eukaryotic and prokaryotic cells of the mammalian intestinal ecosystem. However, the regulatory pathways involved in goblet cell-induced mucus secretion remain largely unknown. Here we demonstrate that the NLRP6 inflammasome, a recently described regulator of colonic microbiota composition and bio-geographical distribution, is a critical orchestrator of goblet cell mucin granule exocytosis. NLRP6 deficiency leads to defective autophagy in goblet cells and abrogated mucin secretion into the large intestinal lumen. Consequently, NLRP6 inflammasome-deficient mice are unable to clear enteric pathogens from the mucosal surface, rendering them highly susceptible to persistent infection. This study identifies the first innate immune regulatory pathway governing goblet cell mucus secretion, linking non-hematopoietic inflammasome signaling to autophagy and highlighting the goblet cell as a critical innate immune player in the control of intestinal host-microbial mutualism.
NLRP6; inflammasome; goblet cell; mucin; autophagy; Citrobacter infection
Tested and effective approaches are available to prevent mental, emotional, and behavioral problems in youth, but such approaches are underused. Communities That Care (CTC) is a coalition-based strategy that aims to increase the use of tested and effective programs by combining the use of scientific evidence and stakeholder consensus to support the community adoption of a scientific approach to preventing mental, emotional, and behavioral problems in youth. A community-randomized trial of CTC was conducted with a sample of 24 communities matched in pairs and assigned randomly to a control or an intervention condition. The findings demonstrate that CTC significantly increases the community-wide adoption of a science-based approach to prevention. Using a meta-analysis technique, this study shows that despite uniformly high-fidelity implementation of CTC in intervention communities, the effect of CTC on the adoption of a scientific approach to prevention varies significantly across the 12 community pairs. Understanding the extent of variation in the effect of CTC on adopting a science-based approach to prevention lays a foundation for identifying aspects of coalition structure, functioning, or capacity that not only may help explain variation in adoption, but may in turn be targeted to strengthen the effect of CTC on the adoption of a science-based approach to prevention within communities.
Communities That Care; coalition; intervention research; prevention; evidence-based practice
To test whether the Communities That Care (CTC) prevention system reduced levels of risk and adolescent problem behaviors community-wide 6 years after installation of CTC and 1 year after study-provided resources ended.
A community-randomized trial.
24 small towns in 7 states, matched within state, randomly assigned to control or intervention condition in 2003.
A panel of 4407 fifth-grade students was surveyed annually through tenth grade from 2004-2009.
A coalition of community stakeholders received training and technical assistance to install CTC, used epidemiologic data to identify elevated risk factors and depressed protective factors in the community, and implemented programs to address their community’s elevated risks from a menu of tested and effective programs for youths aged 10 to 14, their families, and schools.
Levels of risk and incidence and prevalence of tobacco, alcohol, and other drug use, delinquency, and violent behavior by grade 10.
Mean levels of targeted risks increased less rapidly between grades 5 and 10 in CTC than in control communities and were significantly lower in CTC than control communities in grade 10. The incidence of delinquent behavior, alcohol use, and cigarette use, and the prevalence of current cigarette use and past-year delinquent and violent behavior were significantly lower in CTC than in control communities in grade 10.
Using the CTC system can produce enduring reductions in community-wide levels of risk factors and problem behaviors among adolescents beyond the years of supported implementation, potentially contributing to long-term public health benefits.
Rapid molecular typing methods are important tools in surveillance and outbreak investigations of human Salmonella infections. Here we described the development of a three-genes PCR-RFLP typing method for the differentiation of Salmonella species, subspecies and serovars using the Agilent 2100 Bioanalyzer. The fliC, gnd, and mutS genes were PCR-amplified in 160 Salmonella strains representing the two Salmonella species, six subspecies, and 41 different serovars of S. enterica subspecies enterica. PCR products were individually cut with two different restriction enzymes and the resulting 930 restriction patterns were collected using the Agilent 2100 Bioanalyzer followed by cluster analysis. Both species of Salmonella were differentiated by conventional PCR. All of S. bongori tested were gnd PCR negative due to a mismatch at the 3′-end in one the PCR primers. Salmonella subspecies were differentiated into third-teen homogeneous groups representing each of the six subspecies by cluster analysis of restriction patterns generated from the mutS gene cut with AciI. S. enterica subspecies enterica serovars were further differentiated by the combination of the three target genes and five out the six sets of restriction patterns with a discriminatory power of 0.9725 by cluster analysis. The combined RFLP results of five sets of restriction patterns allowed us to assign each of the 160 strains to one of 128 restriction types. During inoculation studies we were able to identify S. Saintpaul and Typhimurium from 24 h pre-enrichment samples using the described method. The use of fliC, gnd, and mutS PCR-RFLP with the Agilent 2100 Bioanalyzer can provide an accessible and automated alternative method for differentiation of Salmonella pathogens.
Salmonella enterica; Bioanalyzer; PCR-RFLP; restriction type; reference collection
Biological information processing is often carried out by complex networks of interconnected dynamical units. A basic question about such networks is that of reliability: if the same signal is presented many times with the network in different initial states, will the system entrain to the signal in a repeatable way? Reliability is of particular interest in neuroscience, where large, complex networks of excitatory and inhibitory cells are ubiquitous. These networks are known to autonomously produce strongly chaotic dynamics — an obvious threat to reliability. Here, we show that such chaos persists in the presence of weak and strong stimuli, but that even in the presence of chaos, intermittent periods of highly reliable spiking often coexist with unreliable activity. We elucidate the local dynamical mechanisms involved in this intermittent reliability, and investigate the relationship between this phenomenon and certain time-dependent attractors arising from the dynamics. A conclusion is that chaotic dynamics do not have to be an obstacle to precise spike responses, a fact with implications for signal coding in large networks.
Vibrio parahaemolyticus is the leading cause of food-borne illnesses associated with the consumption of raw shellfish worldwide. Here, we report 45 draft genomes of V. parahaemolyticus. Thirty-five of them are strains that were isolated from clinical cases in the state of Maryland from 2010 to 2013. The remaining 10 strains were historical isolates, isolated mostly from the West Coast of the United States during the period of 1988 to 2004. The availability of these genomes will allow for future phylogenetic analyses with other V. parahaemolyticus strains.
Shiga toxin-producing Escherichia coli (STEC) strains (n = 194) representing 43 serotypes and E. coli K-12 were examined for clustered regularly interspaced short palindromic repeat (CRISPR) arrays to study genetic relatedness among STEC serotypes. A subset of the strains (n = 81) was further analyzed for subtype I-E cas and virulence genes to determine a possible association of CRISPR elements with potential virulence. Four types of CRISPR arrays were identified. CRISPR1 and CRISPR2 were present in all strains tested; 1 strain also had both CRISPR3 and CRISPR4, whereas 193 strains displayed a short, combined array, CRISPR3-4. A total of 3,353 spacers were identified, representing 528 distinct spacers. The average length of a spacer was 32 bp. Approximately one-half of the spacers (54%) were unique and found mostly in strains of less common serotypes. Overall, CRISPR spacer contents correlated well with STEC serotypes, and identical arrays were shared between strains with the same H type (O26:H11, O103:H11, and O111:H11). There was no association identified between the presence of subtype I-E cas and virulence genes, but the total number of spacers had a negative correlation with potential pathogenicity (P < 0.05). Fewer spacers were found in strains that had a greater probability of causing outbreaks and disease than in those with lower virulence potential (P < 0.05). The relationship between the CRISPR-cas system and potential virulence needs to be determined on a broader scale, and the biological link will need to be established.
The synthesis of ribosomes is an essential process, which is aided by a variety of transacting factors in bacteria. Among these is a group of GTPases essential for bacterial viability and emerging as promising targets for new antibacterial agents. Herein, we describe a robust high-throughput screening process for inhibitors of one such GTPase, the Escherichia coli EngA protein. The primary screen employed an assay of phosphate production in 384-well density. Reaction conditions were chosen to maximize sensitivity for the discovery of competitive inhibitors while maintaining a strong signal amplitude and low noise. In a pilot screen of 31,800 chemical compounds, 44 active compounds were identified. Further, we describe the elimination of non-specific inhibitors that were detergent-sensitive or reactive as well as those that interfered with the high-throughput phosphate assay. Four inhibitors survived these common counter-screens for non-specificity but these chemicals were also inhibitors of the unrelated enzyme dihydrofolate reductase, suggesting that they too were promiscuously active. The high-throughput screen of the EngA protein described here provides a meticulous pilot study in the search for specific inhibitors of GTPases involved in ribosome biogenesis.
PMID: 23606650 CAMSID: cams4460
EngA; GTPase; ribosome biogenesis; enzyme screen
For Salmonella enterica serovar Enteritidis, 85% of isolates can be classified into 5 pulsed-field gel electrophoresis (PFGE) types. However, PFGE has limited discriminatory power for outbreak detection. Although whole-genome sequencing has been found to improve discrimination of outbreak clusters, whether this procedure can be used in real-time in a public health laboratory is not known. Therefore, we conducted a retrospective and prospective analysis. The retrospective study investigated isolates from 1 confirmed outbreak. Additional cases could be attributed to the outbreak strain on the basis of whole-genome data. The prospective study included 58 isolates obtained in 2012, including isolates from 1 epidemiologically defined outbreak. Whole-genome sequencing identified additional isolates that could be attributed to the outbreak, but which differed from the outbreak-associated PFGE type. Additional putative outbreak clusters were detected in the retrospective and prospective analyses. This study demonstrates the practicality of implementing this approach for outbreak surveillance in a state public health laboratory.
Salmonella enterica serovar Enteritidis; bacteria; high-throughput nucleotide sequencing; whole-genome sequencing; pulsed-field gel electrophoresis; infectious disease outbreaks; public health laboratory surveillance
A conference entitled ‘Human microbiome science: Vision for the future’ was organized in Bethesda, MD from July 24 to 26, 2013. The event brought together experts in the field of human microbiome research and aimed at providing a comprehensive overview of the state of microbiome research, but more importantly to identify and discuss gaps, challenges and opportunities in this nascent field. This report summarizes the presentations but also describes what is needed for human microbiome research to move forward and deliver medical translational applications.
In 2012 we conducted an integrated ecological assessment of the marine environment of the Pitcairn Islands, which are four of the most remote islands in the world. The islands and atolls (Ducie, Henderson, Oeno, and Pitcairn) are situated in the central South Pacific, halfway between New Zealand and South America. We surveyed algae, corals, mobile invertebrates, and fishes at 97 sites between 5 and 30 m depth, and found 51 new records for algae, 23 for corals, and 15 for fishes. The structure of the ecological communities was correlated with age, isolation, and geomorphology of the four islands. Coral and algal assemblages were significantly different among islands with Ducie having the highest coral cover (56%) and Pitcairn dominated by erect macroalgae (42%). Fish biomass was dominated by top predators at Ducie (62% of total fish biomass) and at Henderson (35%). Herbivorous fishes dominated at Pitcairn, while Oeno showed a balanced fish trophic structure. We found high levels of regional endemism in the fish assemblages across the islands (45%), with the highest level observed at Ducie (56% by number). We conducted the first surveys of the deep habitats around the Pitcairn Islands using drop-cameras at 21 sites from depths of 78 to 1,585 m. We observed 57 fish species from the drop-cams, including rare species such as the false catshark (Pseudotriakis microdon) and several new undescribed species. In addition, we made observations of typically shallow reef sharks and other reef fishes at depths down to 300 m. Our findings highlight the uniqueness and high biodiversity value of the Pitcairn Islands as one of the least impacted in the Pacific, and suggest the need for immediate protection.
Next-generation sequencing is being evaluated for use with food-borne illness investigations, especially when the outbreak strains produce patterns that cannot be discriminated from non-outbreak strains using conventional procedures. Here we report complete genome assemblies of two Salmonella enterica serovar Heidelberg strains with a common pulsed-field gel electrophoresis pattern isolated during an outbreak investigation.
The analysis of stimulus/response patterns using information theoretic approaches requires the full probability distribution of stimuli and response. Recent progress in using information-based tools to understand circuit function has advanced understanding of neural coding at the single cell and population level. In advances over traditional reverse correlation approaches, the determination of receptive fields using information as a metric has allowed novel insights into stimulus representation and transformation. The application of maximum entropy methods to population codes has opened a rich exploration of the internal structure of these codes, revealing stimulus-driven functional connectivity. We speculate about the prospects and limitations of information as a general tool for dissecting neural circuits and relating their structure and function.
Here, we report draft genome sequences of 26 isolates of Salmonella enterica subsp. enterica, representing eight serotypes, which were isolated from cows in a Pennsylvania dairy herd, the farm on which they were reared, and the associated off-site heifer-raising facility over an 8-year sampling period.